Monoclonal antibodies were introduced in 1975 by Kohler and Milstein. The concept implies fusing immune lymphocytes with a continuous cell line, for example myeloma. A cloning and selection method makes it possible to select and culture cells producing a specific antibody. This cell clone then originates from one original cell ("monoclonal") and produces exactly identical copies of a specific antibody. These monoclonal antibodies have been prepared against a long line of molecules, and this type of antibody has been used and is still being used to an immense extent. Also the commercial development has been considerable, and today a large number of monoclonal antibodies are being marketed, above all for diagnostic purposes.
This development of mouse-monoclonal antibodies (i.e. prepared by means of immune mouse lymphocytes) has not been followed up with human monoclonal antibodies, in spite of the extensive extensive predicted therapeutic use of these antibodies. This is due to the fact that it is extremely difficult to produce, in a practically feasible manner, immune human lymphocytes for immortalization by cell hybridization or transfection. Ethically, patients, volunteers etc. cannot today be immunized with the molecules against which it is desired to produce human monoclonal antibodies, i.e. tumor-associated antigens, bacterial and viral antigens, toxins etc. Up to the present, the procedure was to seek out patients suffering from, for example, infections, tumors etc., thereby to gain access to immune lymphocytes so-called in vivo sensitized lymphocytes). From the practical point of view, this technique is unacceptable.
The development therefore has occurred within so-called in vitro immunization, implying that non-immune human lymphocytes have been immunized in cell culture environment,i thereby to ensure that all types of antigens can be used. These in vitro immunized human lymphocytes are then fused with myeloma or lymphoblastoid cells for continuous production of human monoclonal antibodies. Alternatively, the immune lymphocytes can be transfected with a suitable viral or bacterial genome, thereby to immortalize the cells.
In other words, in vitro immunization is the only technique by which human monoclonal antibodies can be prepared in the future in a practically feasible and ethically acceptable manner. The technique has been developed during a number of years in the murine system, and there exist today, for mouse cells, in vitro immunization methods which function well (Borrebaeck, 1986).
In the human system to which considerable resources have been made available in order to provide for in vitro immunization, there is today nothing which corresponds to the mouse system. This is because the activation requirements for human cells are partly different, and above all because peripheral blood lymphocytes have been used which, possibly, exist in a "deeper" state of rest. Still there are some reports on human in vitro immunization against a few types of haptens, for example bombesin (Ho et al. 1985) and DNP-HSA (Teng et al. 1985), and red corpuscles (Strike et al. 1984; Hoffman & Hirst 1985). These in vitro immunization systems have no lowest common denominator, and different techniques have been used with very different results, frequently with very low yields of specific hybridomas. In addition, there is at present no system at all capable of producing a satisfactory primary immunological response in vitro against thymus dependent antigens. Using different types of T-cell derived lymphokines is one technique which has functioned well in connection with mouse cells (Borrebaeck & Moller 1986) but which alone is not sufficient in the human system. Other attack techniques that have been tried in order to support a human in vitro immunization for the production of human monoclonal antibodies comprise using
(1) adjuvant peptides, such as muramyl dipeptide; PA1 (2) monokine supplemented medium; PA1 (3) separation of cell populations with Sepharose/Sephadex, gelatin, plastic, nylon wool adherence, antibody spanning, complement lysis, affinity column; PA1 (4) polyclonal activators, such as endotoxins (LPS), lectins (PHA, PWM, Con A), Staphylococcus aureus cells, protein A or G; PA1 (5) special sera, such, as ABO, FCS, rabbit serum.
It has now been found that the problem primarily lies int. al. in the removal of cell subpopulations (possibly cytotoxic) capable of suppressing or otherwise preventing the antigen-specific immunological response in vitro.